pLenti-NOTCH2-sgRNA

Cat. No.: P748602

Specifications

Product Name

pLenti-NOTCH2-sgRNA

Product Line

Biologische Produkte

Lagerung und Versand

Storage

Store at -80°C

Verschickt in

Dry ice packs + Cold packs

Stabilität und Lagerung

Stored at -20 ℃, valid for at least 6 months;Stored at -80 ℃, valid for at least one year.

Application Protocols

Tested applications, recommended MOI/dose, transduction enhancers, antibiotic selection, and assay timelines are not specified for this item; refer to CoA/Spec Sheet and institutional SOPs.

General guidance (non-protocol):

• Include appropriate positive/negative controls (non-targeting sgRNA, known pathway sgRNAs) and technical replicates.
• Confirm delivery and expression by the available marker system if present (e.g., fluorescence or antibiotic resistance), noting that specific markers for this SKU are not specified.
• Validate genome editing or transcriptional modulation outcomes with orthogonal assays (sequencing, qPCR of downstream targets, functional readouts).

Biological Roles

This reagent targets the NOTCH2 gene via CRISPR sgRNA. The following summarizes the biological context of NOTCH2 (literature/general background, not product-specific):

• NOTCH2 is one of four mammalian Notch receptors (NOTCH1–4) mediating juxtacrine signaling that regulates cell fate decisions, differentiation, and tissue homeostasis.
• Activation mechanism: Ligand (e.g., Jagged/Delta-like) binding triggers ADAM metalloprotease cleavage followed by gamma-secretase–mediated intramembrane proteolysis, releasing the NOTCH2 intracellular domain (NICD). NICD translocates to the nucleus and modulates transcription with CSL/RBPJ and co-activators.
• Functional roles: Influences lineage specification in hematopoietic, neural, and epithelial contexts; modulates proliferation and differentiation in a context-dependent manner.
• Gene structure: Large single-pass transmembrane receptor with multiple EGF-like repeats extracellularly and RAM/ANK domains intracellularly; numerous context-specific target genes are downstream via CSL sites.

Implication for experiments:

• sgRNA-mediated perturbation of NOTCH2 allows dissection of Notch pathway contributions to specific phenotypes. Given paralog redundancy and crosstalk, include appropriate pathway controls (e.g., NOTCH1/3 sgRNAs, gamma-secretase inhibition) and pathway readouts (HES/HEY transcriptional reporters) to interpret outcomes.

Note: Specific guide sequence, target exon, and species specificity for this SKU are not specified; consult the CoA/Spec Sheet.

Buffer Applications

This product is not a buffering agent and is not used to prepare analytical or biological buffer systems. For experimental use, follow standard cell culture or molecular biology buffer practices compatible with the supplied formulation. Any buffer composition associated with this SKU is not specified; refer to CoA/Spec Sheet.

Green Alternatives

While “green chemistry” solvent substitutions are not directly applicable to a lentiviral sgRNA reagent, researchers can consider greener/safer alternatives in genome editing workflows to reduce hazardous waste and biosafety burden.

• Alternative delivery formats (general, literature):
  • RNP delivery (Cas9 protein + synthetic sgRNA): Avoids use of viral vectors, reduces integration risk, and limits off-target exposure time; reduced biological waste handling compared to lentiviral methods.
  • Non-viral plasmid transfection (electroporation/lipid): Eliminates viral packaging and associated disinfectant usage; suitable for many immortalized cell lines.
  • AAV vectors: Non-enveloped, different biosafety profile; selection depends on cell type and payload size (tradeoffs in packaging capacity and tropism).

Comparison (conceptual):

• Lentiviral sgRNA
  • Pros: Broad tropism with VSV-G, stable expression via integration, effective for hard-to-transfect cells.
  • Cons: BSL-2 handling, bleach-based decontamination, potential integration-related complexities.
• RNP
  • Pros: Transient activity, minimal genomic footprint, often high efficiency; reduced biosafety waste.
  • Cons: Requires protein and guide preparation; transient expression may limit certain long-term studies.
• Plasmid transfection
  • Pros: Simple, scalable, no viral work.
  • Cons: Lower efficiency in some primary cells; transient unless selection/integration systems are used.

Pharmaceutical Uses

This is a research-use-only lentiviral/plasmid sgRNA reagent and is not intended for human or veterinary use, clinical diagnostics, or therapeutic manufacturing.

General formulation context (non-clinical):

• In research settings, lentiviral sgRNA constructs are formulated in sterile aqueous buffers for in vitro cell culture applications. Excipients, stabilizers, and residual solvents (if any) are application- and supplier-specific and are not specified for this item.
• No pharmacopeial monograph applies to engineered lentiviral sgRNA constructs; regulatory-grade release testing (sterility, endotoxin, residuals) is not implied unless explicitly documented. For this SKU, such attributes are not specified; refer to CoA/Spec Sheet.

Physical Properties

• Physical state/appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Buffer composition: Not specified for this item; refer to CoA/Spec Sheet.
• Concentration/titer: Not specified for this item; refer to CoA/Spec Sheet.
• pH, osmolarity: Not specified for this item; refer to CoA/Spec Sheet.
• Classical physicochemical constants (bp, mp, density, refractive index, logP): Not applicable to nucleic acid/viral vector reagents.

General/literature context (for planning only):

• Plasmid DNA preparations and lentiviral stocks are aqueous and do not have meaningful boiling/melting points like small molecules; stability is primarily governed by nuclease avoidance, buffer composition, and freeze–thaw history.
• Typical storage for viral stocks is ≤ -80°C to preserve functional titer; repeated freeze–thaw cycles reduce activity (literature).
• Viscosity and surface adsorption can affect handling of low-titer viral preparations; low-binding plastics are commonly used (literature).

Quality and Grades

• Grade/purity: Not specified for this item; refer to CoA/Spec Sheet.
• Lot-specific data (sequence, sterility, endotoxin, titer): Not specified for this item; refer to CoA/Spec Sheet.

Interpretation and typical expectations (general guidance):

• Sequence identity: sgRNA constructs are commonly verified by Sanger sequencing of the guide insert and scaffold junctions; full plasmid verification may be provided for some lots (not specified here).
• Functional quality for lentiviral stocks is commonly reported as transducing units per mL (TU/mL) or genome copies per mL (GC/mL); exact assay used and thresholds are lot-specific (not specified here).
• Sterility and mycoplasma testing may be performed for biological reagents intended for cell culture; methods and acceptance criteria vary by supplier (not specified here).
• Endotoxin levels are relevant for primary cells and in vivo research; thresholds are application-dependent (not specified here).

How to use the information: Consult the CoA/Spec Sheet for this SKU for confirmation of backbone identity, guide sequence(s), selection markers, promoter architecture, and any release specifications that affect your assay (e.g., titer for MOI calculations).

Reaction and Applications

As a pLenti vector encoding an sgRNA targeting NOTCH2, this reagent is intended for genome engineering workflows rather than small-molecule reactions.

Typical research applications (general, literature-based):

• CRISPR-Cas9 knockout: Delivery of NOTCH2-targeting sgRNA into cells that stably express Cas9 or via co-delivery with Cas9 enables generation of indels at the NOTCH2 locus, facilitating gene disruption and loss-of-function studies.
• CRISPR interference/activation (CRISPRi/a): When used with dCas9-KRAB or dCas9-activator fusions, an sgRNA targeting promoter or enhancer elements of NOTCH2 can repress or upregulate transcription (guide position/context dependent; guide design for this SKU is not specified).
• Pooled screening: Libraries often use lentiviral sgRNA delivery for pooled genetic screens; a NOTCH2-directed construct can serve as a positive control or a focused perturbation in signaling studies.
• Stable cell line generation: Lentiviral delivery supports integration and stable expression of sgRNA in dividing and non-dividing cells (subject to vector design; specifics for this SKU are not specified).

Practical considerations (non-protocol):

• Match the sgRNA with the appropriate Cas nuclease (e.g., SpCas9 requires NGG PAM in the genomic target). The exact protospacer and PAM context for this item are not specified; verify with sequence documentation.
• Validate editing by targeted sequencing or genotyping assays and confirm on-target effects with orthogonal readouts.
• Use appropriate controls (non-targeting sgRNA, sgRNA to a safe-harbor locus) to interpret phenotypes.

Reaction Conditions

Classical reaction conditions (solvent, temperature, catalysts, yields) do not apply to this biological reagent.

General experimental considerations for lentiviral sgRNA usage (non-protocol and not product-specific):

• Matching systems: Ensure compatibility between sgRNA scaffold and your Cas effector (e.g., SpCas9 vs SaCas9). The sgRNA scaffold identity for this SKU is not specified.
• Cell delivery: Method (transduction vs transfection), cell density, and exposure time are key determinants of success and vary by cell type. Specific conditions for this SKU are not provided.
• Selection and timing: If the construct includes a selectable marker or reporter, optimize selection windows and assay timing for on-target editing to manifest; details for this SKU are not specified.
• Validation: Use targeted deep sequencing or mismatch-cleavage assays to quantify editing frequency and confirm on-target activity; include controls to assess off-target effects.

For actionable, stepwise conditions, refer to institutional SOPs and the item’s CoA/Spec Sheet. This section intentionally avoids procedural detail in the absence of product-specific documentation.

Safety and Handling

• GHS classification, signal word, H-statements, pictograms: Not specified for this item; refer to SDS.
• Biosafety considerations (general guidance): pLenti-class vectors and lentiviral particles are typically handled under BSL-2 containment with appropriate institutional approvals. Follow institutional biosafety committee (IBC) policies for lentiviral work, including engineering controls (biosafety cabinet), training, and waste decontamination.
• PPE: Lab coat, disposable nitrile gloves, and safety glasses at minimum; use additional PPE (face protection, sleeve covers) per institutional biosafety risk assessment.
• Incompatibilities and deactivation: Avoid exposure to nucleases and extremes of pH/temperature. For spill response and decontamination of lentiviral materials, freshly prepared 0.5–1% sodium hypochlorite or appropriate virucidal disinfectants are commonly used per institutional SOPs (consult SDS/IBC guidance).
• Aerosol minimization: Use sealed rotors for centrifugation and practice gentle pipetting to minimize aerosol generation when handling viral stocks (general biosafety best practice).
• First aid (overview): In case of exposure, follow institutional exposure control plan; wash affected skin with soap and water, flush eyes with water for at least 15 minutes, and seek medical evaluation per workplace policy. Defer to SDS for authoritative instructions.

Note: This product is for research use only.

Solvent Selection

This product is an aqueous biological reagent; classical solvent selection for organic reactions is not applicable.

• Working medium: Typically handled in sterile aqueous buffers compatible with mammalian cell culture. Exact buffer for this SKU is not specified; refer to CoA/Spec Sheet.
• Compatibility considerations (general):
  • Avoid solvents or additives that inactivate viral particles (e.g., strong detergents, alcohols at high percentages) when working with infectious stocks.
  • Maintain isotonicity and physiological pH when applying to cells.
• If downstream cloning or plasmid amplification is intended instead of viral delivery, standard molecular biology buffers apply (TE, nuclease-free water), chosen to balance DNA stability and downstream enzyme compatibility.

Storage and Reconstitution

• Storage conditions (from Product Data): Store at -80°C.
• Shipping (from Product Data): Dry ice packs + Cold packs.
• Thawing/handling (general best practices):
  • Keep on ice during handling and minimize time at ambient temperature.
  • Avoid repeated freeze–thaw cycles; prepare single-use aliquots upon first thaw to preserve functional integrity.
  • Use low-retention, nuclease-free plasticware; maintain aseptic technique for cell culture applications.
• Buffer composition, stabilizers, and recommended diluent: Not specified for this item; refer to CoA/Spec Sheet.
• Reconstitution: Not specified for this item; refer to CoA/Spec Sheet. If provided lyophilized (not specified), reconstitute with nuclease-free buffer per documentation.
• Long-term stability/shelf life: Not specified for this item; refer to CoA/Spec Sheet.

Note: For research use only. Always consult the item-specific CoA/Spec Sheet and SDS for definitive handling and storage information.

Structure and Identity

This product is a lentiviral vector/plasmid construct designed to express a CRISPR single guide RNA (sgRNA) targeting NOTCH2.

• Product name indicates: pLenti-NOTCH2-sgRNA (lentiviral format expressing an sgRNA against the NOTCH2 gene)
• Molecular formula: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular weight: Not specified for this item; refer to CoA/Spec Sheet.
• SMILES: Not applicable for nucleic acid/viral vector reagents.
• InChIKey: Not applicable for nucleic acid/viral vector reagents.
• CAS/CID: Not applicable/not assigned for this engineered research reagent.
• Structural features (general):
  • Backbone: pLenti-class vector (literature: lentiviral transfer plasmid enabling packaging into VSV-G or other envelopes; exact backbone features for this SKU are not specified).
  • Cargo: sgRNA expression cassette targeting the NOTCH2 locus (guide sequence, promoter, and scaffold details are not specified for this item).
  • Format: Provided as a biological reagent suitable for -80°C storage; exact concentration, buffer, and titer are not specified.

2D description (general): Circular plasmid map typically includes an origin of replication, an antibiotic resistance marker, and an sgRNA cassette; when packaged, the functional unit is a pseudotyped lentiviral particle carrying the sgRNA expression cassette. Specific map, sizes, and selectable markers are not specified for this item; refer to CoA/Spec Sheet.

Synthetic Utility

In place of classical synthetic chemistry utility, this reagent provides synthetic biology utility within genome engineering workflows.

• Functional role: Supplies an sgRNA expression cassette targeting NOTCH2 to program CRISPR effectors (e.g., Cas9, dCas9-KRAB/activators) toward a defined genomic locus.
• Retrosynthetic perspective (molecular biology): NOTCH2 perturbation can be incorporated into modular constructs, enabling combinatorial studies (e.g., dual-sgRNA cassettes for multiplex editing) or pathway epistasis mapping.
• Cloning interoperability (general): pLenti backbones often support Golden Gate, Gibson, or ligase-based cloning for guide insertion; exact cloning sites for this SKU are not specified. Researchers typically verify insert orientation and sequence by Sanger sequencing before packaging.
• Downstream assays: Enables loss-of-function phenotyping, transcriptomic profiling (RNA-seq to capture pathway changes), and functional reporter assays. Always include off-target assessment appropriate to your tolerance for risk and phenotype criticality.

Item-specific details such as backbone map, promoters, selection markers, and cloning sites are not specified; consult the CoA/Spec Sheet before planning cloning or packaging steps.

Target Specificity

• Intended target: NOTCH2 gene (as indicated by product name pLenti-NOTCH2-sgRNA).
• Species, guide sequence, PAM context, target exon/intron, and any off-target assessment: Not specified for this item; refer to CoA/Spec Sheet.
• Construct format and selectable markers (if any): Not specified for this item; refer to CoA/Spec Sheet.

Note: For CRISPR applications, specificity depends on the exact 20-nt protospacer and genomic context. Users should confirm sequence details and validate on-target editing in their model system.